Both genetics and environment are critical risk factors in the epidemic of type 2 diabetes. The genetic basis is exemplified by the high degree of concordance of type 2 diabetes in identical twins, the strong influence of family history on the risk of developing diabetes, and the high incidence of diabetes in certain ethnic populations. In addition, criticalenvironmental factors have been identified, including obesity, inactivity, a suboptimal intrauterine environment, aging, and, at a cellular level, chronic hyperinsulinemia, hyperglycemia, and hyperlipidemia, among others. Since both genotype and cellular environment converge to influence cellular function at the level of gene and protein expression, we hypothesize that alterations in gene expression in nondiabetic individual sat high risk for developing diabetes ("prediabetes') mediate this risk. [unreadable] [unreadable] Thus, we propose to utilize high density oligonucleotidearrays to study differential gene expression in tissue biopsy samples from metabolically characterized human subjects at high risk for the development of diabetes. These studies should permit us to identify novel genes and expressed sequence tags (ESTs), which are responsible for the development of diabetes. We have chosen to focus on "prediabetes," or the time period prior to the onset of clinical disease, in order to define early, and potentially pathogenic, alterations in gene expression before they are obscured by secondary changes resulting from hyperglycemia and other metabolic consequences of overt disease. Thus, we will direct our efforts on nondiabetic subject groups defined by specific riskfactors, including (1) genetic background (positive or negative family history of diabetes, stratified by current metabolic profile), (2) prenatal environment (low birth weight, stratified by current metabolic profile), (3) postnatal environment, assessed by differential expression in monozygotic twins discordant for obesity and/or insulin sensitivity. In parallel, we will begin functional characterization of genes confirmed to be differentially expressed as a function of diabetes risk in multiple risk subgroups, by using small interfering RNA (siRNA) to inactivate selected genes in cell culture models of insulin action. These studies should help to define pathophysiology of diabetes and ultimately help to develop more effective and specific methods of prevention and treatment of type 2 diabetes.